The Interaction of Viperin with Hepatitis C Virus Non-Structural Protein 5A Inhibits the Catalytic Activity of Viperin

The radical SAM enzyme viperin exerts a wide range of antiviral effects through both the synthesis of the antiviral nucleotide 39-deoxy-39, 49-didehydro-CTP (ddhCTP) and through its interactions with various cellular and viral proteins. Here we investigate the interaction of viperin with hepatitis C virus non-structural protein 5A (NS5A) and the host sterol regulatory protein, vesicle-associated membrane protein A (VAP-33). NS5A and VAP-33 form part of the viral replication complex that is essential for copying the RNA genome of the virus. Using transfected enzymes in HEK293T cells, we show that viperin binds to both NS5A and VAP-33 independently and that this interaction is dependent on all three proteins being localized to the ER membrane. Co-expression of viperin with VAP-33 and NS5A was found to reduce NS5A levels, most likely by increasing the rate of proteasomal degradation. However, co-expression of viperin with VAP-33 and NS5A also reduces the specific activity of viperin by ≈3-fold. This observation suggests that NS5A may have evolved to bind viperin as a strategy to reduce ddhCTP synthesis, thereby reducing possibility of the replication complex introducing this chain-terminating nucleotide during genome synthesis.